1. Field of the Invention
The present invention relates to the isolation of a nucleic acid sequences that encode an enzyme that catalyzes the transfer of electrons to protons for the production of molecular hydrogen, and more particularly to iron hydrogenase and genes encoding for the iron hydrogenase in microscopic organisms known as unicellular green algae.
2. Prior Art
Molecular hydrogen is currently being considered as a candidate for replacing or supplementing fossil fuels and as a source of clean energy. A potential method for producing hydrogen on a commercial scale is the photobiological production of hydrogen by eukaryotic organisms. Green algae respond to anaerobic stress by switching the oxidative pathway to a fermentative metabolism. The fermentation of organic compounds and residual photosynthetic electron trasport in the green algae are associated with hydrogen evolution. The key enzyme hydrogenase, which is synthesized only after an anaerobic adaptation, catalyzes the reversible reduction of protons to molecular hydrogen. This method is capable of generating renewable hydrogen fuel from light and water, which are among nature's most plentiful resources.
The ability of green algae, such as Chlamydomonas reinhardtii, to produce hydrogen from water has been recognized for over 55 years. This reaction is catalyzed by a reversible hydrogenase, an enzyme that is induced in the cells after exposure to a short period of anaerobiosis. However, the activity of the hydrogenase is rapidly lost when cells are illuminated because of the immediate inactivation of the reversible hydrogenase by photosynthetically generated O2.
Methods have been devised to circumvent the hydrogenase inactivation problem. U.S. Pat. No. 4,532,210 discloses the biological production of hydrogen in an algal culture using an alternating light and dark cycle. The process comprises alternating a step for cultivating the alga in water under aerobic conditions in the presence of light to accumulate photosynthetic products (starch) in the alga, and a step for cultivating the alga in water under microaerobic conditions in the dark to decompose the accumulated material by photosynthesis to evolve hydrogen. This method uses a nitrogen gas purge technique to remove oxygen from the culture.
U.S. Pat. No. 4,442,211 discloses that the efficiency of a process for producing hydrogen, by subjecting algae in an aqueous phase to light irradiation, is increased by culturing algae which has been bleached during a first period of irradiation in a culture medium in an aerobic atmosphere until it has regained color and then subjecting this algae to a second period of irradiation wherein hydrogen is produced at an enhanced rate. A reaction cell is used wherein light irradiates the culture in an environment which is substantially free of CO2 and atmospheric O2. This environment is maintained by passing an inert gas (e.g. helium) through the cell to remove all hydrogen and oxygen generated by the splitting of water molecules in the aqueous medium. Although continuous purging of H2-producing cultures with inert gases has allowed for the sustained production of H2, such purging is expensive and impractical for large-scale mass cultures of algae. In view of the foregoing, there remains a need for a microorganism that produces a hydrogenase enzyme suitable for use in a sustainable process of photosynthetic hydrogen production.